This is a revised grant application from a new investigator. The goal of this proposal is to study the mechanisms by which A3 adenosine receptors (A3AR) are cardioprotective for ischemia, and to study the effect of cardiac-specific overexpression of A3 adenosine receptors on resistance to ischemic injury and development of adenosine tolerance. The hypothesis is that A3 adenosine receptors provide cardioprotection via ATP-dependent potassium channels (KATP), involving the sequential activation of Gi/o proteins, PKC, and MAPK pathways. Using A3 adenosine receptor stimulated adult rabbit myocytes, measurements will be made of PKC isoforms, and activation of P44, P42, P38, and JNK will be examined. Specific PKC isoforms and MAPK pathways will be blocked, and the effect on cardioprotection will be assessed in vitro with ischemic myocytes, and in vivo with infarcted rabbit hearts. These same models will be used with pertussis toxin or with specific KATP channel inhibitors to study the role of Gi/o proteins and KATP channels. The second hypothesis is that cardiac A3 adenosine receptor overexpression in transgenic mice will protect from ischemia and prevent adenosine tolerance. Initially, cardiac function will be studied via physiology, biochemistry, and pathology studies. Then resistance to ischemic injury and to adenosine tolerance will be evaluated in a murine infarction model and an isolated heart model of global ischemia. The results will provide understanding of the mechanisms of A3 adenosine receptor cardioprotection, and may allow for development of molecular strategies to treat ischemic heart disease with increased expression of A3 adenosine receptors.